Embryo-initiated oviductal transport in mares.

The hypothesis that equine embryos initiate oviductal transport in mares was tested by placing day 6 uterine embryos in the oviducts of day 2 (n = 10) or day 5 (n = 10) recipient mares and attempting to collect the embryos from the uterus 48 h later. To determine whether the surgical transfer procedure initiated oviductal transport, medium alone was placed in the oviducts of day 2 (n = 10) inseminated mares (sham transfer), and uterine embryo collections were attempted 48 h later. Embryos were transported through the oviduct of day 2 recipients by day 4 (instead of day 5 to 6) in six of ten mares, which was not significantly less (P greater than 0.1) than in day 5 recipients (9 of 10). Oviductal transport was not primarily initiated by the surgical transfer procedure, since oviductal transport occurred in only one sham transfer. There was no significant difference (P greater than 0.1) in the diameter of embryos placed in the oviducts of day 2 and day 5 recipient mares (180 +/- 13.8 versus 187 +/- 11.3 microns, respectively). However, embryos collected from the uterus were significantly smaller (P less than 0.05) in day 2 than in day 5 recipients (375 +/- 85.4 versus 659 +/- 43.6 microns, respectively). One uterine embryo had shed its zona pellucida before being placed in, and transported through, the oviduct of the recipient mare.     

Successful transfer of the embryos of Przewalski's horses (Equus przewalskii) and Grant's zebra (E. burchelli) to domestic mares (E. caballus)

Blastocysts were collected non-surgically from 2 Przewalski's horse and 2 Grant's zebra mares and transferred extra-specifically to domestic horse and donkey recipients. Nine Przewalski's horse embryos were transferred surgically, and 2 non-surgically, to domestic Welsh-type pony mares. After surgical transfer, 7 (77.8%) pregnancies were established and 4 foals were born. Twelve Grant's zebra embryos were transferred surgically to 5 pony and 7 domestic donkey recipients respectively and 1 non-surgically to a donkey; 3 (60%) zebra-in-horse pregnancies were established and 2 went to term. Only 2 (28.6%) zebra-in-donkey pregnancies were established but neither went to term, although one zebra foal was aborted alive at Day 292 but failed to survive. No pregnancies resulted from the non-surgical transfers. Measurement of chorionic gonadotrophin concentrations and parental-specific lymphocytotoxic antibodies in the serum of the recipient animals indicated a pronounced maternal immunological response to the extra-specific embryo, but this could not be correlated with success or failure of pregnancy. The results indicate that extra-specific embryo transfer may be a useful aid to breeding exotic equids in captivity.     

Survival of equine embryos co-cultured with equine oviductal epithelium from the four- to eight-cell to the blastocyst stage after transfer to synchronous recipient mares

In this study we examined the ability of equine oviductal epithelial cells (OEC) to support the development of four- to eight-cell equine embryos in vitro and investigated the ability of co-cultured embryos to continue normal development after transfer to synchronous recipient mares. Equine embryos obtained at Day 2 after ovulation were cultured with or without OEC for 5 days. Those OEC co-cultured embryos that reached the blastocyst stage and embryos recovered from the uterus at Day 7 were surgically transferred to synchronous recipient mares. Co-culture with OEC improved (P < 0.01) development of four- to eight-cell embryos to blastocysts compared to medium alone (11/15 vs 0/6) during 5 days in vitro. Embryos co-cultured with OEC were smaller (P < 0.05) and more delayed in development than Day-7 uterine blastocysts. There was no difference in the Day-30 survival rate of co-cultured blastocysts (3/8) or Day-7 uterine blastocysts (5/8) after transfer to recipient mares. These results indicate that co-culture with OEC can support development of four- to eight-cell equine embryos in vitro and that co-cultured embryos can continue normal development after transfer to recipient mares.     

Production of piglets from in vitro-produced embryos following non-surgical transfer

The objective of this study was to enhance procedures for producing piglets derived from in vitro-produced (IVP) pig embryos by non-surgical embryo transfer (ET). The effects of insertion length for the catheter, asynchrony between the age of donor IVP blastocysts and the recipient estrous cycle, and volume of transfer medium were investigated. The IVP blastocysts at 5 days after in vitro fertilization were placed into porcine zygote medium (PZM)-5 supplemented with 10% (v/v) fetal bovine serum (PZM+FBS) in a 0.25 mL plastic straw (21-40 blastocysts per straw) and then transferred into one uterine horn of recipients using the Takumi(?) catheter for deep intrauterine insertion. Successful production of piglets derived from IVP embryos was achieved following non-surgical ET when the catheter was inserted at more than 30 cm anterior to the spiral guide spirette. The efficiency of piglet production (percentage number of piglet(s) born based on the number of embryos transferred) was greater (P<0.05) in recipients whose estrous cycle was asynchronous to that of donors with a 1-day delay (8.3%) than in those with a 2-day (1.5%) or 3-day (0.9%) delay, while pregnancy and farrowing rates (10-40%) did not differ among treatments. When blastocysts were transferred into recipients with 1.0 or 2.5 mL PZM+FBS, there were no significant differences in farrowing rate (30-40%) or average litter size (4.5-6.7) between treatments. The results of the present study indicate that the insertion length of the deep intrauterine catheter and the degree of asynchrony between donor embryos and recipient estrous cycle influenced on pregnancy and birth outcome following non-surgical transfer of IVP blastocysts.     

Deep freezing of horse embryos

Fourteen horse embryos recovered non-surgically on Days 6-8 after ovulation (Day 0) were cooled slowly to - 35 degrees C (7 embryos) or - 40 degrees C (7 embryos) and stored in liquid nitrogen (- 196 degrees C) for 4-98 days. Surgical transfer of the thawed embryos to unmated recipient mares that had ovulated - 2 to + 1 days with respect to the embryo donors resulted initially in the establishment of 4 conceptuses. However, only one mare maintained her pregnancy to term.     

Measurement of early pregnancy factor activity for monitoring the viability of the equine embryo

The viability of embryos before flushing from donor mares (n = 5) and after transfer to recipient mares (n = 7) was monitored in mare serum by detecting early pregnancy factor (EPF) using the rosette inhibition test (RIT). The EPF activity was measured in donor mares before and after natural mating at natural estrus; after ovulation on Days 2, 5 and 8; and after embryo flushing (Day 8) on Days 8, 9, 10 and 13 after ovulation. The collected embryos were transferred immediately after flushing. The EPF activity in recipient mares were measured on the day of transfer and after embryo transfer on Days 1, 2, 3 and 5. Pregnancy was confirmed on Day 12 to 14 after embryo transfer. The mean EPF activity of donor mares was increased to the pregnant level (> an RI titer score of 10) on Day 2 after ovulation. Two days after flushing the embryos, the EPF activity of donor mares had decreased to the nonpregnant level. Among the 7 recipient mares, 3 mares were diagnosed pregnant on Day 12 after embryo transfer with ultrasound. The EPF activity of the pregnant recipient mares was increased above the minimum level observed in pregnant mares on Days 2 to 3 after transfer. However, among the nonpregnant recipient mares after embryo transfer, the EPF activity of 3 mares remained at the pregnant level only 2 to 3 d and then declined to the nonpregnant level. In one recipient mare, EPF activity did not reach the pregnant level throughout the sample collection. The results of this study indicated that equine EPF can be detected in serum of pregnant mares as early as Day 2 after ovulation. From our observation, we conclude that the measurement of EPF activity is useful for monitoring the in vivo viability of equine embryos and early detection of embryonic death.     

Non-surgical transfer of deep-frozen bovine embryos

Preservation of cattle embryos by methods of deep-freezing has recently been established (1, 11, 12) and provides a valuable addition to the possibilities of controlled breeding by embryo transfer in cattle.Already long distance transport of frozen embryos has been demonstrated (2, 6) and adopted by some commercial interests. However, in all publications to date, embryos have been transferred to recipients by surgical methods, even though non-surgical methods of embryo recovery and transfer would be preferred for commercial embryo transfer.The purpose of the present experiments was to utilize non-surgical methods of embryo recovery and transfer of deep-frozen cattle embryos to demonstrate the feasibility of the procedure for a farm service to interested breeders. The particular advantage of non-surgical embryo transfer methods is that neither the donor nor recipient need to leave the farm. Embryo preservation by freezing obviates the necessity for synchronization of recipients for immediate transfer from the donor and allows considerable freedom in the choice of the recipients and the timing of embryo transfers.     

Postfixation embryo reduction in unilateral and bilateral twins in mares

Multiple ovulations were induced with a pituitary extract in mares, and the development of multiple conceptuses was monitored daily by ultrasound on days 11 to 40. The incidence of abortion (loss of all embryos) was not significantly different between mares with multiple embryos (5 38 mares; 13%) and mares with singletons (4 36 ; 11%). Embryo reduction (elimination of excess embryos) was not detected during the embryo mobility phase (days 11-15) or on the day of fixation of embryos (day 16) in any of 38 mares with multiple embryos. The incidence of postfixation embryo reduction for mares with twins was 64% (18 28 ); however, the incidence for unilateral twins (17 19 ; 89%) was greater (P<0.01) than for bilateral twins (1 9 ; 11%). Reduction of unilateral twin embryos seemed to occur earlier (53% before day 20 and 82% before day 30) than for the set of bilateral twins (day 36). The remaining embryo in all mares in which embryo reduction occurred seemed normal in size and appearance on the last day of examination. However, in four of eighteen mares in which unilateral reduction occurred, the umbilical cord of the remaining embryo was attached in the ventral hemisphere of the all antochorion. This apparent disorientation was not seen in any of 16 bilaterally located embryos or in 16 singletons.     

Production of identical twins by bisection of blastocysts in the cow

Day-8 embryos were recovered by a non-surgical method from superovulated crossbred heifers. Normal expanded blastocysts with a distinct inner cell mass and a trophoblast were released from the zona pellucida and bisected along a sagittal plane into two 'half' blastocysts. Each 'half' blastocyst was replaced in an empty zona pellucida and cultured for 2 h in B2 medium. After culture the 'half' blastocysts were directly transferred to recipient heifers via the cervix. From 11 blastocysts, 11 monozygotic 'half' blastocyst pairs were transferred to 11 recipients: 8 recipients became pregnant, 4 carried twins and one delivered a normal calf and an acardiacus amorphus monster consisting of disorganized embryonic tissues. A further 11 'half' blastocysts were transferred as singletons to 11 recipients. Five recipients were apparently pregnant at Day 42. One returned to oestrus at Day 45, 3 were carrying normal fetuses and 1 a pair of normal twin fetuses when slaughtered at Day 128. It is concluded that even after the first irreversible cellular differentiation which occurs at the blastocyst stage it is still possible to produce identical cattle twins by bisection of the Day-8 blastocyst.     

Ultrasound-guided non-surgical embryo collection in the common marmoset

Experimental primate embryology has been hampered by limited access to embryos. In addition to surgical techniques, the less stressful non-surgical technique of uterine flushing has been developed but has had only limitedly used in recovering pre-implantation embryos from marmoset monkeys. In this study, we introduce the use of ultrasonography during marmoset non-surgical uterine flushing to make the cannulation easier, to further reduce stress, and to ensure thorough uterine flushing. We were able to cannulate in 99% of the transcervical cannulation attempts, repeat the flushing up to 17 times with the same animal, and recover up to 90% of the ovulation products. We also found that 8-cell or earlier stage embryos could be frequently obtained by non-surgical uterine flushing at 4 or 5 days after ovulation. The easiness and effectiveness of this novel ultrasound-guided technique will enable more research groups to study marmoset embryology and facilitate progress in this field.     

Effect of repeated flushing and a prostaglandin analogue on the estrous cycle of pony mares

An experiment was conducted to test the effect of repeated transcervical (non-surgical) uterine flushing and a prostaglandin analogue (PG) on the estrous cycle of pony mares. Uteri in group A were trancervically flushed for embryos 7 to 9 days post ovulation. In addition, group B mares were given 5 ml of PG by intramuscular injection on the day of flushing. Group C served as controls and were not flushed or given PG but were allowed to cycle normally. All mares (except controls) were bred A.I. every other day during estrus. There was no effect on embryo recovery rate from repeated flushing or PG administration. The number of days in estrus was greater for groups A and B than for group C (P<0.05). Length of diestrus was longer for group C than for the other two groups. The total estrous cycle length was similar for all three groups (P>0.05).     

Effect of GnRH treatment during the anovulatory season on multiple ovulation rate and on follicular development during the ensuing pregnancy in mares

Seasonally anovulatory mares were injected, i.m., twice daily with a GnRH analogue (GnRH-A), and hCG was given when the largest follicle reached 35 mm in diameter. In Exp. 1, treatment was initiated on 23 December when the largest follicle per mare was less than or equal to 17 mm. An ovulatory response (ovulation within 21 days) occurred in 17 of 30 (57%) GnRH-A-treated mares on a mean of 15.8 days. The shortest interval to ovulation in control mares (N = 10) was 57 days. The diameter of the largest follicle first increased significantly 6 days after start of treatment. In Exp. 2, treatment was begun on 15 January and mares were categorized according to the largest follicle at start of treatment. The proportion of mares ovulating within 21 days increased significantly according to initial diameter of largest follicle (less than or equal to 15 mm, 9/25 mares ovulated; 15-19 mm, 13/21; 20-24 mm, 20/24; greater than 25 mm, 3/3). The multiple ovulation rate was greater (P less than 0.01) for treated mares (27/86 mares had multiple ovulations) than for control mares (2/35). Treated mares in which the largest follicle at start of treatment was greater than or equal to 25 mm had a higher (P less than 0.01) multiple ovulation rate (9/14) than did mares in which the largest follicle was less than 25 mm (18/72). The pregnancy rate for single ovulators was not different between control mares (26/30 pregnant mares) and treated mares (43/54).(ABSTRACT TRUNCATED AT 250 WORDS)     

Infection of embryos following insemination of donor mares with equine arteritis virus infective semen

The objective was to evaluate the potential risks associated with embryo transfer from mares bred with equine arteritis virus (EAV) infective semen. Twenty-six mares were embryo donors, whereas 18 unvaccinated and EAV antibody seronegative mares were embryo recipients. Of the 26 donor mares, 15 were unvaccinated and seronegative for antibodies to EAV and 11 were vaccinated for the first time with a commercially available modified live virus vaccine against EVA before breeding and subsequent embryo transfer. All donor mares were bred with EAV-infective semen from a stallion persistently infected with the virus. Twenty-four embryos were recovered 7 d post-ovulation; all were subjected in sequential order to five washes in embryo flush medium, two trypsin treatments, and five additional washes in embryo flush medium (prior to transfer). Twelve and seven embryos (Grades 1 or 2) were transferred from the non-vaccinated and vaccinated donors, respectively, and pregnancy was established in 3 of 12 and 2 of 7. Perhaps trypsin reduced embryo viability and pregnancy rate. The uterine flush fluid of 11 mares (9 of 15 and 2 of 11 from non-vaccinated and vaccinated donor groups, respectively) was positive for EAV by VI (confirmed by real-time RT-PCR); the wash fluid from the embryos of nine of these mares was negative following 10 washes and two trypsin treatments. However, the embryo wash fluid from two mares was still positive for EAV after all 10 washes and the two trypsin treatments, and one embryo was positive for EAV. Two of 18 recipient mares had seroconverted to EAV 28 d after embryo transfer. Virus was not detected in any fetal tissues or fluids harvested after pregnancies were terminated (60 d). In conclusion, we inferred that the washing protocol of 10 washes and two trypsin treatments did not eliminate EAV from all embryos; due to limitations in experimental design, this requires confirmation. Furthermore, there may be a risk of EAV transmission associated with in vivo embryo transfer from a donor mare inseminated with EAV infective semen.     

Factors affecting the success of surgical and nonsurgical equine embryo transfer

Nonsurgical embryo recovery was attempted from light-horse and draft mares. Embryo recovery rates were not affected (P>.05) by technician or stallion but were lower (P<.05) from draft mares (44%) than light-horse mares (67%). Sham transfer of embryos on day 8 post-ovulation did not (P>.05) increase the number of mares returning to estrus by 22 days post-ovulation. Method of embryo transfer greatly affected pregnancy rates. Embryos transferred surgically during March–June resulted in 0 of 12 pregnancies versus 13 of 25 pregnancies obtained during July–September, This strongly suggests a seasonal influence on pregnancy rates. Technician influenced (P<.05) the success of nonsurgical transfer (46.2% vs. 7.7%). In addition, protection of the insemination rod with a sheath (guarded method) appeared to provide some advantage over an unguarded method of nonsurgical transfer (54% vs. 23%). Lastly, a preliminary experiment was conducted to evaluate transfer of embryos via flank incision. Four of 5 embryos transferred by this method resulted in a pregnancy at 50 days post estrus.     

Methanol as a cryoprotectant for equine embryos

Equine embryos (n=43) were recovered nonsurgically 7-8 days after ovulation and randomly assigned to be cryopreserved in one of two cryoprotectants: 48% (15M) methanol (n=22) or 10% (136 M) glycerol (n=21). Embryos (300-1000 microm) were measured at five intervals after exposure to glycerol (0, 2, 5, 10 and 15 min) or methanol (0, 15, 35, 75 and 10 min) to determine changes (%) in diameter over time (+/-S.D.). Embryos were loaded into 0.25-ml plastic straws, sealed, placed in a programmable cell freezer and cooled from room temperature (22 degrees C) to -6 degrees C. Straws were then seeded, held at -6 degrees C for 10 min and then cooled to -33 degrees C before being plunged into liquid nitrogen. Two or three embryos within a treatment group were thawed and assigned to be either cultured for 12 h prior to transfer or immediately nonsurgically transferred to a single mare. Embryo diameter decreased in all embryos upon initial exposure to cryoprotectant. Embryos in methanol shrank and recovered slightly to 76+/-8 % of their original diameter; however, embryos in glycerol continued to shrink, reaching 57+/-6 % of their original diameter prior to cryopreservation. Survival rates of embryos through Day 16 of pregnancy were 38 and 23%, respectively (P>0.05) for embryos cryopreserved in the presence of glycerol or methanol. There was no difference in pregnancy rates of mares receiving embryos that were cultured prior to transfer or not cultured (P>0.05). Preliminary experiments indicated that 48% methanol was not toxic to fresh equine embryos but methanol provided no advantage over glycerol as a cryoprotectant for equine blastocysts.     

Effect of embryo age and recipient asynchrony on pregnancy rates in a commercial equine embryo transfer program

In the present study, 809 uterine flushes and 454 embryo transfers performed in mares over a 4-yr interval were examined to evaluate the effects of: (1) the day of embryo collection on recovery rates; (2) the degree of synchrony between donor and recipient mares on pregnancy rates; (3) the recipient day post ovulation on pregnancy rates; and (4) the age of the embryo at recovery on pregnancy rates at 60 days. Uterine flushes were performed on Days 6, 7, 8, 9, and 10 (Day 0 = ovulation) and embryos were transferred to recipients with degrees of synchrony varying between +1 to −6 (recipient ovulated 1 day before through 6 days after the donor). Recipient mares ranged from 2 to 8 days post ovulation. Embryo recovery rates were similar for flushes performed on Day 7 (61%), Day 8 (66%), Day 9 (59%), and Day 10 (56%), but the embryo recovery rate was lower (P < 0.03) for flushes performed on Day 6 (42%) compared with all other days. Pregnancy rates for various degrees of synchrony were as follows: +1 (71%), 0 (77%), −1 (68%), −2 (63%), −3 (66%), −4 (76%), −5 (61%), and −6 (27%). The −6 day of degree of synchrony had the lowest (P < 0.05) pregnancy rate compared with all other days, but there was no significant difference among +1 to −5 days. There was a lower (P < 0.05) pregnancy rate for embryos transferred to recipient mares on Day 2 (33%) compared with mares on Day 3 (66%), Day 4 (66%), Day 5 (62%), Day 6 (55%), Day 7 (58%), and Day 8 (56%). Pregnancy rate was higher (P < 0.05) for Day 7 (76%) embryos compared with Day 6 (50%), Day 8 (64%), and Day 9 (44%) embryos; Day 9 embryos resulted in lower (P < 0.05) pregnancy rates than Days 7 or 8 embryos. In conclusion, this study demonstrated that: (1) embryo recovery rates between Days 7 and 10 were similar and acceptable (e.g., 63% 488/771); (2) the degree of synchrony between donor and recipient mares does not need to be as restricted as previously reported in horses. Acceptable pregnancy rates (e.g., 70%, 99/142) were obtained even when recipient mares ovulated 4 to 5 days after the donors; (3) similar pregnancy rates were obtained when recipient mares received embryos within a large range of days post ovulation (Days 3 to 8); and (4) Day 7 embryos produced higher pregnancy rates when compared with Days 8 and 9 embryos. In clinical terms, the application of these new findings will be beneficial to large equine embryo transfer operations in producing more pregnancies per season.     

Regression and resurgence of the CL following PGF2alpha treatment 3 days after ovulation in mares

The present study was designed to characterize and compare the physiology and ultrasonographic morphology of the corpus luteum (CL) during regression and resurgence following a single dose of native prostaglandin F2alpha (PGF) given 3 days after ovulation, with a more conventional treatment given 10 days after ovulation. On the day of pre-treatment ovulation (Day 0), horse mares were randomly assigned to receive PGF (Lutalyse; 10 mg/mare, i.m.) on Day 3 (17 mares) or Day 10 (17 mares). Beginning on either Days 3 or 10, follicle and CL data and blood samples were collected daily until post-treatment ovulation. Functional and structural regression of the CL in response to PGF treatment were similar in both the Day 3 and 10 groups, as indicated by an abrupt decrease in circulating concentrations of progesterone, decrease in luteal gland diameter and increase in luteal tissue echogenicity. As a result, the mean +/- S.E.M. interovulatory interval was shorter (P < 0.0001) in the Day 3 group (13.2 +/- 0.9 days) than in the Day 10 group (19.2 +/- 0.7 days). Within the Day 3 group, functional resurgence of the CL was detected in 75% of the mares (12 of 16) beginning 3 days after PGF treatment, as indicated by transient major (6 mares) and minor (6 mares) increases (P < 0.05 and < 0.1, respectively) in progesterone. Correspondingly, mean length of the interovulatory interval was longer (P < 0.03) in mares with major resurgence (15.8 +/- 1.6 days) than in mares with minor (11.2 +/- 1.2 days) and no resurgences (13.5 +/- 0.3 days) in progesterone. Structural resurgence of the CL in the Day 3 group and functional and structural resurgence in the Day 10 group were not detected. In conclusion, PGF treatment 3 days after ovulation resulted in structural and functional regression of the CL and hastened the interval to the next ovulation, despite post-treatment resurgences in progesterone.     

Effect of exogenous progesterone on pregnancy rates after surgical embryo transfer in mares

A completely randomized experimental design was used to investigate the effect of supplemental progesterone on pregnancy rates of recipient mares. Every other recipient mare received daily 200 mg progesterone in oil beginning the day of surgical embryo transfer and lasting until either Day 120 of pregnancy or until pregnancy failure was confirmed by ultrasound. Progesterone supplementation did not affect pregnancy rate (P > 0.05). Overall, embryos that did not result in pregnancy were of greater mean diameter than embryos that resulted in pregnancy (P < 0.05). Pregnancy rates tended (P < 0.1) to be greater in recipients that were detected to be ovulating the same day or prior to that of the donor and that had been supplemented with progesterone (75 %) as opposed to untreated control mares of the same synchrony group (40 %). Progesterone supplementation did not affect the incidence of embryonic loss; however, there was a slightly higher loss of pregnancies between Day 15 and 30 in treated versus untreated recipients. There was no effect (P > 0.05) of treatment on pregnancy rate for embryos recovered from fertile versus subfertile donor mares. However, overall, there tended (P < 0.1) to be fewer pregnancies with embryos recovered from subfertile (50 %) as compared to fertile donors (75 %). It was concluded that supplemental progesterone at the dosage and frequency described was not beneficial in improving pregnancy rates in cyclic recipient mares after surgical embryo transfer.     

Effects of recombinant human follicle stimulating hormone on follicle development and ovulation in the mare

The only gonadotrophin preparation shown to stimulate commercially useful multiple ovulation in mares is equine pituitary extract (EPE); even then, the low and inconsistent ovulatory response has been ascribed to the variable, but high, LH content. This study investigated the effects of an LH-free FSH preparation, recombinant human follicle stimulating hormone (rhFSH), on follicle development, ovulation and embryo production in mares. Five mares were treated twice-daily with 450 i.u. rhFSH starting on day 6 after ovulation, coincident with PGF(2alpha) analogue administration; five control mares were treated similarly but with saline instead of rhFSH. The response was monitored by daily scanning of the mares' ovaries and assay of systemic oestradiol-17beta and progesterone concentrations. When the dominant follicle(s) exceeded 35 mm, ovulation was induced with human chorionic gonadotrophin; embryos were recovered on day 7 after ovulation. After an untreated oestrous cycle to 'wash-out' the rhFSH, the groups were crossed-over and treated twice-daily with 900 i.u. rhFSH, or saline. At the onset of treatment, the largest follicle was <25 mm in all mares, and mares destined for rhFSH treatment had at least as many 10-25 mm follicles as controls. However, neither dose of rhFSH altered the number of days before the dominant follicle(s) reached 35 mm, the number of follicles of any size class (10-25, 25-35, >3 mm) at ovulation induction, the pre- or post-ovulatory oestradiol-17beta or progesterone concentrations, the number of ovulations or the embryo yield. It is concluded that rhFSH, at the doses used, is insufficient to stimulate multiple follicle development in mares.     

Repeat superovulation, non-surgical embryo recovery, and surgical embryo transfer in transgenic dairy goats

We investigated the capability of repeat superovulation and non-surgical embryo retrieval, coupled with surgical embryo transfer, to expedite the production of transgenic progeny from transgenic founder dairy goat does. In addition, we compared embryo yields, number of embryos transferred per recipient, pregnancy rates, and offspring born during both the traditional (September-December) and non-traditional (January-May) breeding seasons. Although there were no significant differences, there were numerically more transferable embryos recovered per flush (3.5+/-0.9 vs. 2.4+/-0.9 embryos; mean+/-S.E.M.) and increases in both the proportion of recipients that were pregnant (83 vs. 69% pregnant) and offspring born from total embryos transferred (67 vs. 53% offspring) during the traditional versus the non-traditional breeding season. The transfer of one, two or three embryos did not significantly affect the proportion of pregnant recipients during either season. However, there was a difference (P<0.05) in the proportion of offspring produced for one versus two embryo transfers (89 vs. 44% offspring, respectively) during the non-traditional breeding season. Overall, 14 transgenic offspring were produced from 54 total offspring born, and the kidding interval was reduced to <3 months for six of the seven transgenic does. In summary, repeat superovulation and non-surgical embryo retrieval, coupled with surgical embryo transfer, expedited the production of progeny from transgenic founder does.